Progressive multifocal leukoencephalopathy (PML) is a deadly brain disease caused by the JC polyomavirus (JCV). PML arises in subjects with immunodeficiency in which specific T cells against JCV are lacking, as seen by the absence of T cell receptors (TCRs) recognizing JCV peptides. It is the goal of this proposal to provide to PML patients the missing anti-viral specificity through a new T cell immunotherapeutic strategy. To achieve this, we will perform ex vivo gene therapy on patients' own T cells to express TCR molecules that recognize JCV-infected cells. These TCRs are in turn genetically fused to molecular domains of T cell signaling molecule(s) to create chimeric immune receptors (CIRs) that regulate the T cell activity. We refer to TCR-based CIRs as "TCRCIR". In this application, we propose a staged development and application of anti-JCV designer T cells with our most advanced 2nd generation (Signal 1 + 2) constructs of CIRs, to take them through preclinical development to actual clinical trials. We propose an associated laboratory research program to monitor these studies, and to develop newer, 3rd generation designer T cells that are IL2 independent. Specific Aims for this four-year clinical development application are: Preclinical: To develop anti-JCV "designer T cells": (1) to clone JCV p36- and plOO-specific TCRs from HLA-A2 + PML survivors; (2) to create and test DNA constructs of JCV-specific TCRCIRs in 2nd generation in human T cells and (3)to create and test DNA constructs of JCV-specific TCRCIRs in 3rd generation form in human T cells; Clinical: (4) to perform Phase I clinical trial of 2nd generation anti-JCV designer T cells. By this plan, it is hoped that designer T cells will survive and expand in JCV + lesions in vivo, and eliminate JCV-infected cells. If functioning as intended, the application of anti-JCV designer T cells will parallel in vivo observations of CD8 + JCV-reactive T cells in PML-survivors and induce a "survivor" phenotype in patients who lack such cells endogenously.